A variety of methods have been proposed which measure specific components in specimens, particularly in living bodies and solutions using an attenuated total reflectance (hereinafter referred to as ATR) measuring instrument.
For example, in Japanese Patent Laid-Open No. 9-113439, there is proposed a method of measuring the blood sugar level using a transparent ATR device 51 having a pair of parallel reflecting surfaces opposing each other in which measurement is made with upper and lower lips 52, as a specimen, brought into tight contact with the ATR device 51, as shown in FIG. 7. According to this method, the measurement of the blood sugar level is made through the following procedures: inserting an ATR device 51 between upper and lower lips, and getting the same to be hold firmly by the lips; entering light into the ATR device 51 so that the light is allowed to undergo total reflection repeatedly at the interface between each reflection surface of the ATR device 51 and the lips 52, as shown by the broken line in FIG. 7; and analyzing the light that oozes out of the ATR device 51. The entire disclosure of Japanese Patent Laid-Open No. 9-113439 is incorporated herein by reference in its entirety.
In BME, Vol. 5, No. 8 (Japan ME Society, 1991), there is proposed a method which measures the blood sugar level, the concentration of ethanol in blood, etc. using an ATR device made up of ZnSe optical crystal etc. In the method, measurements are made through the following procedures: bringing the ATR device into tight contact with lip mucosa; entering a laser light with a wavelength of 9 to 11 microns into the ATR device and allowing the light to undergo multiple reflection inside the ATR device; and analyze the absorbed and scattered light. The entire disclosure of BME, Vol. 5, No. 8 (Japan ME Society, 1991) is incorporated herein by reference in its entirety.
According to this method, concentrations of specific components such as glucose, ethanol and cholesterol can be measured non-invasively and in real time. This method is to apply evanescent light (known as ooze-out light) to a quantitative analysis. Only a very small quantity of the light traveling in the ATR device actually enters lips, and the light having entered the lips is affected by components in the body fluid existing in the lips.
For example, in glucose, its light absorption peaks at a wave number of 1080 cm−1; therefore, when applying light with the above wave number to a living body, the quantity of the light absorption of glucose changes depending on the glucose concentration in the living body. Accordingly, if the quantity of the light returned from the living body is measured, the change in quantity of the light absorption of a component in body fluid with change in the concentration of the component can be detected, in other words, the concentration of the component can be obtained.
When measuring the absorbance of a substance surface with an ATR measuring instrument and calculating the concentration of the same using the measured absorbance, the measuring method shown in FIG. 8 has been commonly used.
First, in the background measuring step, the measurement of background is made by entering light emitted by a light source into the ATR device, carrying out spectrometry of a reference, such as air or deionized water, while keeping the ATR device out of contact with a sample as a subject of measurement, and storing the measured results in a memory (S4). The reasons for the background measurement are to correct the wavelength characteristics of a light source and a photodetector and to ensure an accurate absorbance measurement or concentration calculation even after their characteristics have changed with time.
Then, the sample as a subject of measurement is set so that it comes in contact with the ATR device (S5) and measurement is made for the sample (S6).
Calculation is carried out according to the following equation, Log10 (Ib/Im), where Ib represents a detected signal from the photodetector at the time of background measurement and Im a detected signal from the photodetector at the time of measurement for the sample (S7). The calculated value is commonly referred to as absorbance. Since absorbance correlates with concentration of a specific components in a sample, if a calibration curve of absorbance and concentration is prepared in advance, the concentration of a specific component in the sample can be estimated from the calculated absorbance.
The conventional ATR measuring instruments described above, however, have the following problems.
When making measurement of a sample after a certain length of time has elapsed since the completion of background measurement, the intensity of the light source and the sensitivity of the photodetector have changed delicately, which has made accurate measurement of the sample difficult.
Further, when measuring the spectral characteristics of a sample, in order to make the measurement accurate, it is necessary to set the contact position and the contact area, where the sample and the ATR device are in contact with each other, just the same as those at the time of background measurement. However, such setting has been difficult to perform accurately; as a result, the obtained accuracy of measurement has not been satisfactory. Particularly when the sample has been a living body, it has been difficult to accurately position the sample and the ATR device at the time of measurement.